Journal of Analytical & Bioanalytical Techniques
Open Access

Abstract

Simple, isocratic RP-HPLC method with UV detection for measurement of liposoluble vitamins: retinol, �?±-tocopherol, �?³-tocopherol, retinol-palmitate and �?²-carotene in human plasma was developed. Plasma samples were prepared for analysis by liquid-liquid extraction. Ethanol was used for precipitation of proteins. Further extraction of analytes was performed with n-hexane. Extracts were evaporated to dryness under stream of nitrogen and further reconstituted in ethanol before HPLC analysis. Separation of analytes was performed on Zorbax Eclipse XDB-C18 Rapid Resolution, 4.6x100 mm, 3.5 �?¼m chromatographic column with mobile phase consisting of methanol and ethanol (75:25, v/v). Mobile phase flow rate was 1 mL/min. Column temperature was set on 40�?°C and autosampler temperature on 10�?°C. Total run time was 16 minutes. UV detection of retinol and retinol-palmitate was performed on 325 nm, �?±-tocopherol and �?³-tocopherol at 292 nm and �?² carotene on 450 nm. Tocopherol-acetate was used as internal standard. Calibration curves for all analytes were constructed in following ranges: retinol, �?±-tocopherol and �?³-tocopherol (5â�?�?35 �?¼g/mL), �?²-carotene and retinol-palmitate (10â�?�?35 �?¼g/mL). Developed method was successfully used for routine determination of liposoluble vitamins in plasma samples of healthy, middle-aged volunteers.

Biography

Milkica Crevar Saka�? has completed her PhD at the Department of Medicinal Chemistry, Faculty of Pharmacy, University of Belgrade. She works as Teaching Assistant and Researcher at Department of Medicinal Chemistry.

Email: frodo@pharmacy.bg.ac.rs